Aglaia, is able to sensitize resistant HCC cells to TRAIL-mediated apoptosis. Two HCC cell lines, HepG2 and Huh-7, were treated with rocaglamide and/or TRAIL and the induction of apoptosis and effects on the TRAIL signaling pathway were investigated. The in vivo efficacy of rocaglamide was determined in TRAIL-resistant Huh-7-derived tumor xenografts. Rocaglamide significantly sensitized the TRAIL-resistant HCC cells to apoptosis by TRAIL, which resulted from the rocaglamide-mediated downregulation of cellular FLICE-like inhibitory protein and subsequent caspase-8 activation. Furthermore, rocaglamide markedly inhibited tumor growth from Huh-7 cells propagated in severe combined immunodeficient mice, suggesting that chemosentization also occurred in vivo. These data suggest that rocaglamide acted synergistically with TRAIL against the TRAIL-resistant HCC cells. Thus, it is concluded that rocaglamide as an adjuvant to TRAIL-based therapy may present a promising therapeutic approach for the treatment of HCC.The enhancement of apoptosis is a therapeutic strategy used in the treatment of cancer. Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) is a promising antitumor agent. However, hepatocellular carcinoma (HCC) cells exhibit marked resistance to the induction of cell death by TRAIL. The present study investigated whether rocaglamide, a naturally occurring product isolated from the genus  Hepatocellular carcinoma (HCC) is one of the most common and life-threatening types of human malignant tumor . The inaTumor necrosis factor-related apoptosis-induced ligand (TRAIL), a type II transmembrane protein from the tumor necrosis factor family, triggers the extrinsic pathway of apoptosis by binding to the cell surface receptors death receptor (DR) 4 and 5 . These rAglaia (family Meliaceae) (Rocaglamide is isolated from the genus liaceae) . A numbeliaceae) ,14. Howein vitro. Furthermore, the efficacy of rocaglamide in TRAIL-resistant Huh-7-derived tumor xenografts was determined.The aim of the present study was to investigate whether rocaglamide sensitized resistant HCC cells to TRAIL-induced death, by regulation of caspase-8/c-FLIP 2. Rocaglamide (>98% pure) was procured from Enzo Life Sciences . TRAIL was purchased from PeproTech, Inc.  and all chemicals were purchased from Sigma , unless indicated otherwise.HepG2 and Huh-7 cells were obtained from the Shanghai Cell Collection  and cultured in Dulbecco’s modified Eagle’s medium  supplemented with 10% fetal bovine serum , 2 mM glutamine, 100 U/ml penicillin and 100 μg/ml streptomycin. The cells were cultured at 37°C in 5% COFor the investigation of time-dependence, the cells (70% confluent) were treated with rocaglamide (100 nM) and/or TRAIL (100 ng/ml) for different time periods (0–24 h). The cells were then harvested for cell viability analysis. For the investigation of dose-dependence, the cells (70% confluent) were pretreated with rocaglamide (0–100 nM) for 12 h in DMEM, supplemented with 10% FBS, 2 mM glutamine, 100 U/ml penicillin and 100 μg/ml streptomycin, and were then treated with TRAIL (0–100 ng/ml) for an additional 12 h. To investigate treatment with a combination of rocaglamide and TRAIL, the cells were pretreated with either rocaglamide or dimethyl sulfoxide (DMSO) for 12 h, followed by 12 h incubation in the presence of TRAIL. Subsequently, 24 h after rocaglamide or DMSO treatment, the cells were harvested and the cell lysates were prepared, according to previous methods  and stor4/well) were seeded in 96-well plates in complete culture medium and incubated for 24 h. The cells were then exposed to 100 nM rocaglamide and/or 100 ng/ml TRAIL for 24 h. The control cells were treated with DMSO at a concentration equal to that used for the drug-treated cells. The complete culture medium was then removed and MTT  was added to each well and the plate was incubated for 2 h at 37°C in a humidified incubator. The solution was then removed from the wells and 200 μl DMSO was added to each well prior to agitation. The absorbance at 570 nm was read using a microplate reader . The value for the vehicle-treated cells was considered to indicate 100% viability. Furthermore, a crystal violet assay was carried out. Briefly, the cells (1.0×105/ml) were seeded in a 12-well plate for 12 h, and treated with TRAIL (0–100 ng/ml) and/or RocA(1–100 nM) for 12 h. The treated cells were washed with phosphate-buffered saline (PBS), fixed with 4% paraformaldehyde for 15 min, and stained using crystal violet  for a further 30 min.HepG2 and Huh-7 cells RNA control or siRNA FLIP (Santa Cruz Biotechnology) of high purity were delivered into the HepG2 cells in the 12-well plate using Lipofectamine 2000  according to the manufacturer’s instructions. Briefly, 2 μl Lipofectamine 2000 was added to 50 nmol/l siRNA in a final volume of 100 μl culture medium and the solution was added to the washed cells, which were then incubated at 37°C for 4 h. The transfection mixture was then removed and the cells were incubated with fresh complete medium for an additional 36 h. Finally, the cells were exposed to TRAIL or DMSO for 12 h for subsequent experiments.5) were harvested and resuspended in 400 μl 1× binding buffer with 4 μl FITC annexin V and 4 μl PI. Flow cytometric analysis was then performed using a Becton-Dickinson FACSCalibur™ system; BD Biosciences, Franklin Lakes, NJ, USA) and FlowJo 7.6.3 software .The HepG2 and Huh-7 cells were seeded and cultured in each well of a 12-well plate for 24 h and then pretreated with either rocaglamide or DMSO for 12 h, followed by 12 h incubation in the presence of TRAIL. An annexin V/propidium iodide (PI) apoptosis detection kit  was used for the detection of apoptotic and necrotic cells. The cells (5×106), suspended in 100 μl mix , were implanted subcutaneously into the right flank of 10 female SCID mice (6-week-old) and then randomly divided into two equal groups, one of which received an intraperitoneal injection of rocaglamide  and the other, used as a vehicle control, received olive oil alone (n=5). These treatments were performed once daily for 32 days and the tumor volumes and body weights of the animals were measured twice a week. The tumor volumes (mm3) were calculated using the following formula: Tumor volume = LS2 / 2, where L is the longest diameter and S is the shortest. At the end of the experiments, the mice were sacrificed and tumor samples were harvested, fixed in formalin and embedded in paraffin as tissue sections for immunohistochemical analysis. The animal experiments were performed in accordance with the relevant institutional and national regulations and the research procedures were approved by Tongji Medical College . The SCID mice were purchased from Beijing HFK Bioscience, Co., Ltd. .The Huh-7 cells  according to the manufacturer’s instructions. The primary antibody was omitted in the first step to establish a negative control, which yielded negative results. The percentage of positive cells was independently determined by two examiners. At least five random fields of each section were visualized (magnification ×400) and analyzed using a Nikon OPTIPHOT 150 microscope  connected to a SPOT Insight charge-coupled device camera . The values are expressed as the mean ± standard deviation (SD).Immunohistochemical staining was performed using either 1:1 hematoxylin and 0.5% eosin (H&E) or an anti-cleaved caspase-3 antibody , as previously described . ApoptosWhere indicated, the data are expressed as the mean ± SD from at least three independent experiments. Statistical analysis of the data was conducted using Student’s t-test. P<0.01 was considered to indicate a statistically significant difference.To investigate whether rocaglamide enhanced TRAIL-mediated apoptosis in HCC cells in the present study, HepG2 and Huh-7 cells, which are highly chemoresistant to TRAIL , were seTRAIL-induced apoptosis is mediated by activation of the caspase cascade . In partAs pro-apoptotic B-cell lymphoma 2 (Bcl-2) family proteins, including BH3-interacting domain death agonist (BID), Bcl-2-associated X protein (BAX) and p53-upregulated modulator of apoptosis (PUMA), are able to sensitize cancer cells to TRAIL-induced apoptosis , the prec-FLIP, which is highly homologous to caspase-8 but is catalytically inactive, is able to bind to caspase-8 and the FADD and inhibit TRAIL-induced apoptosis, thus disrupting the death-inducing signaling complex (DISC) . NotablyTRAIL triggers the extrinsic pathway of apoptosis by binding to cognate DR4 or DR5 , which rIn order to determine the effect of downregulation of c-FLIP on the rocaglamide-induced effect, the present study also examined whether the apoptosis-inducing effects of rocaglamide were also exerted in c-FLIP-knockdown HepG2 cells. siRNA was used to specifically downregulate c-FLIP and an siRNA vector served as a control. This treatment resulted in a marked suppression in the levels of c-FLIP of ~70% . The celin vivo effect of rocaglamide and TRAIL on the growth of HCC xenograft tumors. As natural killer cells produce TRAIL in vivo (in vivo study. The Huh-7 cells were subcutaneously injected into the right flanks of SCID mice and, when tumors were visible, the mice were matched for tumor volumes and were assigned to a control group and a rocaglamide-treated group. Tumor volumes in the rocaglamide-treated group were ~45±12% compared with the control group (in vivo.To confirm whether the synergistic effect of rocaglamide and TRAIL in resistant cell lines had potentially relevant clinical implications, the present study investigated the  in vivo ,27, rocaol group . Rocaglaol group , suggestin vivo by examining tumor tissues harvested from the control and rocaglamide-treated mice using H&E, TUNEL and cleaved caspase-3 staining. In the untreated controls, staining with H&E revealed a compact mass of epithelial cells, whereas following rocaglamide treatment, the tumor appearance was that of loose epithelial cell aggregates with increased interspersed mesenchymal cells (in vivo investigation suggests that rocaglamide is an effective drug, which has the potential to inhibit the growth of HCC cell-derived tumors in vivo.The present study further investigated the effect of rocaglamide on apoptosis in vitro and in vivo conditions.HCC is a significant cause of mortality worldwide due to its poor prognosis . The abiAs the type of cancer cell in HCC may affect the response of the cell to therapeutic agents, the robustly TRAIL-resistant HCC cell lines, HepG2 and Huh-7, were selected . FurtherTRAIL-based therapy offers promising therapeutic potential due to its specificity for cancer cells without evident adverse effects on normal cells ,32. Howein vitro. These findings are consistent with previous studies indicating that the activation of caspanse-8 induces apoptosis and sensitizes cancer cells to TRAIL (The detailed mechanisms of resistance to TRAIL in HCC cells remain to be elucidated. The regulation of DISC-engaged molecules, including c-FLIP and caspase-8, has been observed to contribute to the sensitivity of TRAIL-mediated apoptosis in cancer cells ,33,34. Ito TRAIL ,37.c-FLIP inhibits the apoptotic signaling cascade by preventing the recruitment and activation of caspase-8 at the DISC , demonstTargeting the transcriptional activation of c-FLIP is considered to be a promising approach for the downregulation of c-FLIP expression in cancer cells . Howeverin vitro. Furthermore, rocaglamide markedly inhibited the growth of tumors derived from Huh-7 cells in vivo in a xenograft mice model. Thus, these findings provide significant evidence for the development of rocaglamide as a novel therapeutic agent for use as an adjuvant to TRAIL in the treatment of HCC.The present study demonstrated that rocaglamide, a naturally occurring product, sensitized chemoresistant HCC cells to TRAIL-mediated apoptosis by decreasing the expression of c-FLIP and activating caspase-8


The association between heart failure (HF) and herpes zoster has rarely been studied. We investigated the hypothesis that HF may increase the risk of herpes zoster in Taiwan using a nationwide Taiwanese population-based claims database.N = 4785). For a comparison cohort, three age- and gender-matched control patients for every patient in the study cohort were selected using random sampling . All subjects were tracked for 1 year from the date of cohort entry to identify whether or not they had developed herpes zoster. Cox proportional-hazard regressions were performed to evaluate 1-year herpes zoster-free survival rates.Our study cohort consisted of patients who received a diagnosis of HF in 2001 ~ 2009 (p < 0.001) than that of the controls during the 1-year follow-up. Our study also investigated whether HF is a gender-dependent risk factor for herpes zoster. We found that male patients with HF had an increased risk of developing herpes zoster .The main finding of this study was that patients with HF seemed to be at an increased risk of developing herpes zoster. Of the total patients, 211 patients developed herpes zoster during the 1-year follow-up period, among whom 83 were HF patients and 128 were in the comparison cohort. The adjusted hazard ratio (AHR) of herpes zoster in patients with HF was higher : 1.54 ~ 2.78; The findings of our population-based study suggest that patients with HF may have an increased risk of herpes zoster. These health associations should be taken into consideration, and further studies should focused on the cost-effectiveness of the herpes zoster vaccine should be designed for HF patients. Herpes zoster manifests as characteristic painful vesicular skin lesions and related neurological disorders usually unilaterally grouped and limited to 1 to 3 dermatome. Herpes zoster is caused by spontaneous reactivation of a latent varicella-zoster virus (VZV) that resides in sensory ganglia and dorsal nerve roots following a varicella infection . Post-heThe estimated lifetime risk of herpes zoster is 10% ~ 30%, and the incidence and severity markedly increase after the age of 50 years -11. The Heart failure (HF) has high prevalence rate among elderly patients and increases with aging ,29. DissNational Health Insurance (NHI) is a mandatory health insurance program in Taiwan that provides comprehensive coverage for medical care. Up to 99% of the population was enrolled by 2009. All claims data are collected in the NHI Research Database (NHIRD) and are managed by the Taiwan National Health Research Institutes (NHRI).The NHIRD consists of comprehensive healthcare data provided to researchers, including ambulatory care records, inpatient care records, registration files, catastrophic illness files, and various data regarding drug prescriptions of the insured among 23.5 million Taiwanese residents. Data used to perform the analyses conducted in this study were retrieved from the LHID 2005, a subset of the NHIRD. The LHID2005 consists of all the original medical claims for 1,000,000 enrollees' historical ambulatory data and inpatient care data under the Taiwan NHI program from 1997 to 2010, and the database was created and is publicly released to researchers.The NHRI, which constructed and maintains the LHID2005 database, further scrambles this encrypted information before the database is released to researchers. The NHRI reported that there were no statistically significant differences in age or gender between the randomly sampled group and all beneficiaries of the NHI program. To maintain claims data accuracy, the NHI’s routine practice of performing cross-checks and validations of medical claims ensures the accuracy of the NHIRD diagnostic coding.th Revision, Clinical Modification (ICD-9-CM) codes 398.91, 402.01, 402.11, 402.91, 404.01, 404.03, 404.11, 404.13, 404.91, 404.93, or 428) between January 2001 and December 2009. The date of the initial diagnosis of HF was assigned as the index date for each HF patient. To improve data accuracy, the HF selection criterion required that all case ICD-9 codes by assigned by a cardiologist. We also chose selection criteria for herpes zoster patients. We only included herpes zoster cases in this study if they received ≥ 2 herpes zoster diagnoses for ambulatory care visit or ≥ 1 diagnosis for inpatient care.We used a study cohort and a comparison cohort to examine the relationship between HF and herpes zoster. We identified 4785 first-time hospitalizations with a discharge diagnosis of HF  before or after the study period were excluded from both cohorts. We also identified relevant comorbidities, including hypertension (ICD-9-CM 401.X-405.X), diabetes mellitus (ICD-9-CM 250.X), hyperlipidemia (ICD-9-CM 272.X), systemic lupus erythematous (ICD-9-CM 710.0), COPD (ICD-9-CM 491.X), rheumatoid arthritis (ICD-9-CM 714.0), and cancer (ICD-9-CM 140.X-208.X).2), the number of physicians per 100,000 people, the percentage of people with a college education, the percentage of people over 65 years of age, and the percentage of agricultural workers. Because levels 4, 5, 6, and 7 contained few HF cases, they were combined into a single group, thereafter referred to as level 4.For our study of urbanization, all 365 townships in Taiwan were stratified into 7 levels according to the standards established by the Taiwanese NHRI based on a cluster analysis of the 2000 Taiwan census data, with 1 referring to the most urbanized area and 7 referring to the least urbanized. The criteria on which these strata were determined included the population density , whichever came first. After adjusting for urbanization level, monthly income, region, and comorbidities as potential confounders, we performed a Cox proportional-hazards analysis stratified by gender, age group, and index year to examine the risk of herpes zoster during the 1-year follow-up in both cohorts. We further classified the gender and age factors in both groups. Hazard ratios (HRs) and 95% confidence intervals (CIs) were calculated to quantify the risk of herpes zoster. The results of comparisons with a two-sided p value of < 0.05 were considered to represent statistically significant differences.All data processing and statistical analyses were performed with the Statistical Package for Social Science (SPSS) software, vers. 18.0  and SAS vers. 8.2 . Pearson Helsinki Declaration. This study was also evaluated and approved by the Kaohsiung Medical University Hospital Institutional Review Board (KMUH-IRB-EXEMPT-20130059).Insurance reimbursement claims adopted in this study were from Taiwan’s NHIRD, which is available for research purposes. This study was conducted in accordance with the p < 0.001), hyperlipidemia (p < 0.001), diabetes (p < 0.001), and COPD (p < 0.001) were more prevalent in the HF cohort than the comparison cohort. We also found that cases had a greater tendency to have a lower monthly income (p < 0.001), and to reside in southern and eastern Taiwan, and reside in less-urbanized communities (p < 0.001) compared to controls.The research design of this study is shown in Figure p < .001). The overall incidence rate was higher in the HF cohort (17.61 per 1000 patient-years) than in the comparison cohort (8.95 per 1000 patient-years).There were 211 patients diagnosed with herpes zoster during the 1-year follow-up, including 83 HF patients (1.7%) and 128 patients in the comparison cohort (0.9%). The Kaplan-Meier survival curves are shown in Figure p < 0.001) than for comparison patients. After adjusting for potential confounders, our results showed that newly diagnosed HF was associated with an approximately 2.07-times greater risk of herpes zoster , compared to non-HF patients  (Table p < 0.001) had a higher risk of developing herpes zoster than female subjects . For 41 ~ 65 year olds, there was the highest risk  for developing herpes zoster (Table We further investigated whether HF is a time-dependent risk factor for herpes zoster and divided HF patients into 3 groups according to the follow-up period. For the three different follow-up periods, there was still statistical significance between the case and comparison groups. The 1-month follow-up period had the highest statistical significance (AHRs: 4.52; 95% CI: 2.28 ~ 8.96; This cohort study demonstrated that, after controlling for other herpes zoster risk factors, patients with HF were at a 2.07 times increased risk than the general population to experience herpes zoster during the 1-year follow-up period. After further stratification by age, gender, and follow-up period, we found a consistent increased herpes zoster risk in HF patients compared to the general population. To our knowledge, this is the first cohort study to investigate the risk of herpes zoster in patients with HF.Patients with suppressed cell-mediated immunity cause by immunosuppressive disorders or therapies, such as elderly people, patients with autoimmune disease, malignancy, or diabetes, are well described as having a higher risk of developing herpes zoster ,15,19-24The major morbidity caused by herpes zoster is post-herpetic neuralgia, which consumes a lot of medical resources . The paiThe present study has a number of strengths. First, this is a large-scale follow-up study using the well-established nationwide NHI research database. The study cohort was highly representative of the general population. The medical comorbidities is likely complete and accurate because the NHI is a compulsory and universal healthcare system with a very high coverage rate in Taiwan. In addition, while racial differences are considered to be a factor that influences the risk of herpes zoster , approxiNevertheless, some insufficiencies should be addressed. First, almost all herpes zoster and HF cases were diagnosed clinically without serologic confirmation or standardized procedure, so diagnoses based on ICD-9-CM codes may be less accurate. However, the NHI Bureau in Taiwan randomly interviews patients and reviews medical records every year to confirm the validity of the diagnoses and quality of care. In addition, the validity of the coding for herpes zoster and HF in administrative data was shown to be good ,43. SecoIn conclusion, the present study suggests a positive association between HF and herpes zoster. Physicians, including dermatologists and cardiologists, should be aware of the greater incidence of herpes zoster among patients with HF. They should also be familiar with proper antiviral therapy, as well as acute and chronic pain management. Immune-compromised patients should receive the herpes zoster vaccine for preventive purposes. Future studies are needed to investigate the possible mechanism between different stages of HF and herpes zoster.


The data revealed that on an inert substrate the diffusion rate of 18.5-kDa MBP increased in GalC-expressing cells, while the diffusion coefficient of PLP was decreased in sulfatide-containing cells. Similarly, when cells were grown on myelination-promoting laminin-2, the lateral diffusion coefficient of PLP was decreased in sulfatide-containing cells. In contrast, PLP's diffusion rate increased substantially when these cells were grown on myelination-inhibiting fibronectin. Additional biochemical analyses revealed that the observed differences in lateral diffusion coefficients of both proteins can be explained by differences in their biophysical, i.e., galactolipid environment, specifically with regard to their association with lipid rafts. Given the persistence of pathological fibronectin aggregates in multiple sclerosis lesions, this fundamental insight into the nature and dynamics of lipid-protein interactions will be instrumental in developing myelin regenerative strategies.In the central nervous system, lipid-protein interactions are pivotal for myelin maintenance, as these interactions regulate protein transport to the myelin membrane as well as the molecular organization within the sheath. To improve our understanding of the fundamental properties of myelin, we focused here on the lateral membrane organization and dynamics of peripheral membrane protein 18.5-kDa myelin basic protein (MBP) and transmembrane protein proteolipid protein (PLP) as a function of the typical myelin lipids galactosylceramide , and sulfatide, and exogenous factors such as the extracellular matrix proteins laminin-2 and fibronectin, employing an oligodendrocyte cell line, selectively expressing the desired galactolipids. The dynamics of MBP were monitored by  Myelin is produced by oligodendrocytes (OLGs) in the central nervous system (CNS), where it enwraps axons in a multilamellar fashion to enable fast conduction of the action potential in vivoApart from the important functional role of these structural myelin components for myelin assembly, a pivotal role of the surrounding extracellular matrix (ECM) is also apparent. ECM serves as a scaffold, providing signals to maintain and regulate cellular processes such as proliferation, migration and differentiation z-scan point FCS, circular scanning FCS (s-FCS) and RICS to investigate the dynamics of membrane associated PLP and MBP in the presence of GalC and/or sulfatide, which were selectively expressed in oligodendrocyte-derived OLN-93 cells. In addition, we investigated whether Ln2 and Fn could modulate these dynamics and as such interfere with myelin biogenesis and assembly. The data revealed differences in the lateral diffusion coefficients of PLP and 18.5-kDa MBP, as dictated by their biophysical environment, specifically with regard to their association with detergent-resistant lipid rafts, and, in addition, by the nature of the ECM on which the cells were grown.Lipid rafts, commonly characterized by their detergent insolubility, are considered dynamic membrane platforms, playing a role in various cellular processes z-scan FCS and RICS measurements, cells were cultured on 8-well Labtek-I slides , pre-coated with poly-L-lysine , Fn  and Ln2  in phenol-red free DMEM at a cell density 30,000 per well, one day prior to transfection. The cells were transfected with PLP-eGFP plasmid  or 18.5-kDa MBP-eGFP . The production of retroviral particles and the subsequent infection and selection of OLN-93 cells were performed according to Maier et al. The cDNAs encoding ceramide galactosyltransferase (cgt) and galactosylceramide 3′-sulfotransferase (cst) were kind gifts of Drs. Matthias Eckhardt  and Brian Popko , respectively. cgt and cst were cloned into the One day after transfection with PLP-eGFP or 18.5-kDa MBP-eGFP, detergent extract preparation with 20 mM CHAPS and discontinuous OptiPrep density gradient centrifugation were performed as previously described Samples were mixed with reducing sample buffer and heated for 30 min at 37°C. Proteins were separated by 10% SDS-PAGE and subjected to immunoblot analyses as described previously 24 hours after transfection with PLP-eGFP or 18.5-kDa MBP-eGFP, antibody staining of the cell surface lipids GalCer and sulfatide were performed on live cells at 4°C. After blocking non-specific binding with 4% bovine serum albumin in phosphate-buffered saline (PBS), cells were incubated with primary antibody for 30 min, washed three times and incubated for 25 min with TRITC-conjugated antibodies . The cells were fixed with 4% paraformaldehyde (PFA) PBS for 20 min at RT, after which the nuclei were stained with DAPI . O1  and O4 (anti-sulfatide) were both a kind gift of Dr. Guus Wolswijk 2/s at 22°C, diffusion coefficient application note of PicoQuant) with a total laser power of 10 µW before the objective (∼4 µW in solution). All in vivo measurements were performed at room temperature to reduce cell mobility and with an excitation power of 2 µW to minimize bleaching (∼0.8 µW in solution). The diffusion coefficients of both PLP-eGFP and 18.5-kDa MBP-eGFP were determined at the bottom plasma membrane to minimize distortion of the point-spread-function (PSF) of the oil objective. Moreover, in this manner, the effect of the different ECM proteins on the diffusion of the proteins could be studied. The software packages PAM and MIA FCS and RICS measurements were performed on a home-built laser scanning pulsed interleaved excitation fluctuation imaging (PIE-FI) setup as described before z-plane for each experiment, FCS measurements were performed at multiple z-positions. Hereby, the laser light was first focused slightly above the cell membrane and then subsequent point FCS measurements were performed at 10 different planes with a separation of 200 nm between planes, moving downwards  GmbH & Co. KG Karlsruhe, Germany) was used. To minimize axial drift and photobleaching during the measurement, the following scanning procedure was applied: all 10 planes were measured sequentially for 5 s. After completion of one stack, the first position was refocused using a home-built perfect focus system. The procedure was repeated a total of 12 times resulting in an acquisition time of 60 s per plane. From the z-position corresponding to the highest mean intensity . For each bin, an intensity trace and auto-correlation function (ACF) was calculated, and both were averaged over 10 consecutive positions to increase signal-to-noise of the data. Then, points with clearly aberrant intensity or ACF traces, likely caused by vesicles diffusing through the focus, were omitted and all remaining bins of several measurements per cell were averaged  of at least three independent experiments. The statistical significance was calculated by a two-tailed Student′s t-test for comparison between two means and by a one-way ANOVA followed by Newman-Keuls posttest to compare more than two means. A p value of p<0.05 was considered statistically significant.To investigate the effect of the major myelin galactolipids GalC and sulfatide on the dynamics and lateral membrane organization of the myelin-specific proteins MBP and PLP, we took advantage of a rat derived oligodendrocyte cell line, OLN-93. These cells represent immature OLGs and they neither synthesize PLP and MBP nor GalC and sulfatide. Accordingly, this enabled us to apply a bottom-up approach where we first stably overexpressed the enzymes see  responsiAfter having determined the expression and surface localization of GalC and sulfatide in the OLN-G and OLN-GS monoclonal cell lines, we transiently transfected the cells with eGFP-tagged 18.5-kDa MBP. To solely determine the effect of extracellular leaflet localized galactolipids on the intracellular distribution of inner leaflet associated peripheral membrane protein 18.5-kDa MBP, we first examined the effect of OLN-P, OLN-G and OLN-GS on the inert coating material PLL. As shown in In different cell types, including OLGs, GalC and sulfatide are able to integrate into small membrane microdomains known as lipid rafts or detergent-resistant membranes z-stack of images was collected to determine the position of the bottom plasma membrane where the maximum amount of protein was localized  than in OLN-P cells (0.25±0.04 µm2/s) and OLN-GS cells (0.25±0.03 µm2/s)  than the rates observed in OLN-P cells (0.23±0.01 µm2/s) and OLN-GS cells (0.28±0.03 µm2/s)  . To obtant model . Consist3 µm2/s) .z-stacks were taken and the three-dimensional information analyzed. As shown in Next, it was of particular interest, given the uniqueness of the OLN-93 cell system, to investigate whether GalC and/or sulfatide could affect the lateral distribution and dynamics of the integral membrane protein PLP. To obtain detailed information on subcellular localization in a reliable manner, 2/s; α = 0.467) than in GalC-expressing OLN-G cells  or parental OLN-P cells   . Accordiz-scan FCS analysis revealed that no significant differences in the diffusion rates of 18.5-kDa MBP were detected for the different cell types on the different ECM substrates (data not shown). s-FCS measurements showed that the lateral diffusion rate of PLP in OLN-93 cells grown on Ln2 was similar in parental cells (OLN-P) and in GalC-expressing cells, i.e., 0.13±0.01 µm2/s and 0.12±0.02 µm2/s, respectively. However, in the case of OLN-GS cells grown on Ln2, the rate decreased to 0.05±0.01 µm2/s , Fn (0.09±0.01 µm2/s) or Ln2 (0.12±0.02 µm2/s), no significant differences were observed, which further supports the specificity of sulfatide in ECM-dependent PLP processing. Given the observed quantitative differences in lateral diffusion rates of PLP in the presence of different galactolipids as a function of the nature of the ECM, we then further investigated the co-localization degree of PLP with GalC and sulfatide on Ln2 and Fn. When grown on Ln2, the degree of co-localization of PLP with GalC in OLN-G cells (0.77±0.03) was significantly higher than the level of co-localization observed with GalC and sulfatide in OLN-GS cells  associate with detergent-insoluble microdomains. Given the asymmetric distribution of MBP and GalC, this effect presumably relies on a transmembrane GalC-mediated stimulus.Myelin galactolipids GalC and sulfatide are crucial for maintaining myelin integrity as is apparent from knockout experiments of enzymes responsible for their synthesis. In the absence of galactolipids, PLP expression remains unaltered however the protein no longer integrates within CHAPS-resistant microdomains z-scan point FCS and RICS analysis revealed that the mobility of 18.5-kDa MBP was significantly higher in OLN-G cells compared to OLN-GS. The apparent difference in raft association between the peripheral protein MBP (association) and the transmembrane protein PLP (integration) is also reflected by the much slower diffusion rate seen for PLP than for 18.5-kDa MBP in the presence of GalC. Differences in the mode of protein interaction with microdomains have been reported before to give rise to differences in lateral diffusion coefficients in situ that GalC-positive microdomains are largely confined to the myelin membranes, whereas sulfatide containing microdomains are restricted to the cell body and primary processes (unpublished observations). Further investigations will be necessary to understand the role of GalC- and/or sulfatide-enriched microdomains in MBP dynamics in myelin membranes, and may benefit from the application of artificial membranes such as GUVs and Giant Plasma Membrane Spheres Remarkably, although 18.5-kDa MBP is a peripheral membrane protein, its dynamics are affected by the presence of galactolipids, in particular GalC. Thus, 18.5-kDa MBP co-distributed to a major extent with GalC in spite of their asymmetric membrane localization. Moreover, in GalC-expressing cells, a substantial fraction (∼ 30%) of the protein was recovered in CHAPS-insoluble microdomains. This effect was absent in galactolipid deficient cells and, surprisingly, in cells expressing both GalC and sulfatide. Intermixing of sulfatide with GalC pools may repress a specific transmembrane-induced GalC effect on MBP distribution in the inner leaflet and, therefore, it is tempting to suggest that GalC may function as a cell surface transmitter, propagating the signal that triggers MBP′s peripheral association with microdomains. Indeed, GalC-containing liposomes or glycol-nanoparticles containing GalC/sulfatide, when added to primary OLGs caused a redistribution of MBP at the cytoplasmic surface and GalC at the extracellular surface together with a redistribution of some phosphorylated proteins involved in signal transduction It is becoming increasingly apparent that changes in the ECM affect myelination efficiency and alterations in the lateral organization of myelin proteins have been proposed to be part of the underlying mechanism


The required conversion factor varies significantly across phytoplankton taxa and environmental conditions, but little information is available on its response to iron limitation. In this study, we examine the role of iron limitation, and the interacting effects of iron and light availability, on the coupling of photosynthetic electron transport and CO2-assimilation in marine phytoplankton. Our results show that excess irradiance causes increased decoupling of carbon fixation and electron transport, particularly under iron limiting conditions. We observed that reaction center II specific rates of electron transport  increased under iron limitation, and we propose a simple conceptual model for this observation. We also observed a strong correlation between the derived conversion factor and the expression of non-photochemical quenching. Utilizing a dataset from in situ phytoplankton assemblages across a coastal – oceanic transect in the Northeast subarctic Pacific, this relationship was used to predict ETRRCII: CO2-assimilation conversion factors and carbon-based primary productivity from FRRF data, without the need for any additional measurements.Iron availability directly affects photosynthesis and limits phytoplankton growth over vast oceanic regions. For this reason, the availability of iron is a crucial variable to consider in the development of active chlorophyll a fluorescence based estimates of phytoplankton primary productivity. These bio-optical approaches require a conversion factor to derive ecologically-relevant rates of CO On the last day of the experiment, additional samples were collected for pigment analysis by high pressure liquid chromatography (HPLC), and the determination of absorption spectra using the quantitative filter technique (QFT)  samples. Sterile, trace metal clean techniques were used at all times.The oceanic centric diatom ld study ,67. Ironld study , prepareld study . All cululturing . Cultureulturing . Acclimaa] as described below.In addition to the iron addition experiment, seawater samples were collected at five hydrographic stations  spanning a coastal to open ocean transect in the NE subarctic Pacific . Collecta] were filtered onto pre-combusted 25 mm glass fiber filters (GF/F) using low vacuum pressure (<5 mm Hg) and analyzed following the method of Welschmeyer . This result confirms that the initial phytoplankton assemblage was iron-limited (a] in the control treatments is likely attributable to a decrease in grazing pressure and to changes in the light environment (i.e. lower and less fluctuating light). Iron addition also significantly affected phytoplankton photophysiology, as demonstrated by rapid changes in the parameters σPSII and Fv/Fm derived in the dark-regulated state , remained high and relatively constant in the iron-limited control, it declined rapidly after iron addition, and remained ~25% lower than that of the initial phytoplankton assemblage . The stperiment . In contreatment .2-assimilation and ETRRCII to iron addition is further visualized in max), as well as the derived conversion factor Φe:C/nPSII for α and Pmax, throughout the experiment. Values for α and Pmax were derived from the 14C-based and FRRF-based PvsE curves shown in a-normalized CO2-assimilation, ETRRCII or Φe:C/nPSII . Similarly, the Pmax for chl a-normalized CO2-assimilation remained relatively constant in the control, and did not show a statistically significant increase after iron addition   . In contperiment . The obs control . This di control . These rRCII and Φe:C/nPSII observed in our data, we examined changes in additional FRRF-derived ChlF parameters, measured on day 3 after iron addition. We choose day 3 for the in-depth analysis of our data, but trends observed on this day were representative of those observed throughout the experiment. The parameter Fq′/Fv′ represents the efficiency of charge separation in functional RCII (A oxidized) at any given light level, and therefore always equals one at zero irradiance. On day 3 after iron addition, we observed higher Fq′/Fv′ for the iron-limited control at all irradiance levels  is the product of Fq′/Fv′ and Fv′/Fm′ [q′/Fm′ was higher in the iron addition treatment than in the iron-limited control  (Given the similar light and iron-dependent responses of Φtic fit) .vsE curves of CO2-assimilation and ETRRCII in mono-specific laboratory cultures of two open ocean phytoplankton species. The results, summarized in -1) in the low iron cultures were 68% and 49% of iron-replete growth rates in T. oceanica and C. polylepis, respectively . In iron-limited T. oceanica, σPSII increased by 15%, while it increased by 5% in C. polylepis. The iron dependent changes in μ, Fv/Fm and σPSII was statistically significant in both species . Chlorophyll a-normalized CO2-assimilation at Pmax remained relatively constant in both species . In contrast, we observed a 90% increase in ETRRCII at Pmax in T. oceanica under iron-limited growth conditions. C. polylepis also exhibited an increase in ETRRCII at Pmax under iron-limited conditions, but this increase was not statistically significant . Regardless of species-specific differences, both species showed the same trend of increased Φe:C/nPSII and NPQNSV under iron limitation (e:C/nPSII) was increased in the iron addition treatment and the abundance of prymnesiophytes (higher Φe:C/nPSII) was decreased  used to calculate growth rates are given in brackets. Fv/Fm and σPSII are values from cultures in the dark regulated state (10 min of 5 μmol quanta m-2s-1 at 730 nm), measured on the day of CO2-assimilation experiments. The error is SD of 3 biological replicates. Changes in these parameters are statistically significant for T. oceanica  and C. polylepis . Pmax for CO2-assimilation (mol C mol chla-1 s-1) and ETRRCII (mol e- mol RCII-1 s-1) were derived from PvsE curves as described in the methods section. The error is the 95% confidence interval of the Pmax derived from the fit to data from 6 whole curve measurements . The conversion factor Φe:C/nPSII for Pmax was derived as the quotient of Pmax for ETRRCII and Pmax for CO2-assimilation. The error is the propagated error from numerator and denominator. NPQNSV was estimated as Fo′/Fv′ from the last ST acquisition during each light level of the PvsE curves. The values shown are from the first and last step of the PvsE curves (4 and 800 μmol quanta m-2 s-1). Each NPQNSV value is the mean of 2 values measured on 3 biological replicates. Changes in response to iron limitation are statistically significant for both species .2-assimilation and photosynthetic electron transport in natural phytoplankton assemblages and mono-specific laboratory cultures. We show that both of these environmental variables significantly influence Φe:C/nPSII, which has important implications for the use of FRRF measurements to infer rates of CO2-assimilation in oceanic waters. Below, we first discuss the observed increase in Φe:C/nPSII under excess light and low iron conditions in the context of previously reported values. We then discuss the effects of iron and light on phytoplankton photophysiology, and suggest a simple conceptual explanation for the observed increase in ETRRCII under iron limitation. We hypothesize, that iron and light-dependent changes in Φe:C/nPSII are driven by the need to dissipate excess excitation energy, caused by either excess light, or the effects of iron limitation on the ETC. In this context, we discuss the correlation between Φe:C/nPSII and NPQNSV, and examine the potential significance of this finding in the context of marine primary productivity studies.Our results provide new insight into the effects of iron and light availability on the coupling between COe:C/nPSII, derived from our measurements of ETRRCII and CO2-assimilation, varied significantly in response to light and iron availability. In our field experiment, the addition of iron caused the value of Φe:C/nPSII at light saturation (Pmax) to decrease by 66% within 6 days , vs. 3–4 hours in the field and 30 min in the laboratory for 14C-assimilation experiments (light levels applied simultaneously to different samples). As has been shown by Halsey et al. [e:C/nPSII in the iron-limited relative to iron-replete samples. Additionally, the longer incubation time in CO2-assimilation experiments might have exacerbated cumulative processes such as photodamage under excess irradiance. To address this issue, we did not utilize the part of the PvsE curves showing photo-inhibition. However, we cannot rule out any differential cumulative effects of photoinhibition on ETRRCII and 14C-assimilation at Pmax. This could potentially decrease CO2-assimlation at Pmax relative to ETRRCII at Pmax and lead to overestimation of our Φe:C/nPSII values at Pmax. Notwithstanding these potential sources of uncertainty in the absolute value of Φe:C/nPSII, the good agreement between our estimated Φe:C (assuming ~ 500–700 mol chl a mol RCII-1) and those of previous studies suggests that our observations are robust. More importantly, potential offsets in the absolute values of Φe:C/nPSII do not diminish the significance of the relative, iron and light-dependent changes we observed in this parameter (discussed below).Ideally, measurements of ETR:C/nPSII . Howeverstically . Furthery et al. ,17,99 any et al. , the usee:C/nPSII in phytoplankton field assemblages and mono-specific laboratory cultures . Cha. Chae:C/position  reflect RCII and CO2-assimilation can act to increase Φe:C, and therefore the conversion factor Φe:C/nPSII  formed at the end of the ETC can also be used for nitrate and sulphate reduction [RCII from CO2-assimilation, increasing the value of Φe:C. Similarly, before the formation of NADPH, pseudo-cyclic electron flow can reduce O2 and create a water-water cycle of electron transport, also increasing Φe:C .,100.RCIIeduction , photoreeduction , or respeduction . These a:C (e.g. ). Pseudo:C (e.g. ) or afte:C (e.g. ). Cyclic:C  (e.g. [RCII and therefore Φe:C/nPSII (.Iron limitation directly affects the photosynthetic ETC and thereby modulates the light-dependent changes in the conversion factor Φ:C/nPSII . ImportaI) e.g. ,116–118)–118e:C/nI)  (e.g. . Cyclic I)  , and an increase in chl a per functional RCII (1/nPSII) [e:C/nPSII under iron limitation, regardless of light intensity  ,90. This1/nPSII) , and up-1/nPSII) ,90, woulntensity .RCII and CO2-assimilation. This high excitation pressure may be a result of short-term exposure to excess irradiance as well as the effect of iron limitation on the ETC. This purely photophysiological interpretation can be extended to observations made in mixed phytoplankton communities. Here, fluctuating light and low iron conditions will select for species with the best ability to control high excitation pressure by adjusting the flow of excitation energy into, and the flow of electrons out of PSII.In summary, we suggest that it is the effect of high excitation pressure, which causes a de-coupling of ETRRCII increases under iron limitation. This observation may seem counter-intuitive, and it is important to emphasize that our results do not imply an overall increase in photosynthetic electron transport under low iron conditions. Rather, our observations point to an increase in the rate of charge separation at each individual RCII, independent of the reduced total cellular concentration of these RCII. We show that the overall efficiency of PSII photochemistry in the light-regulated state, Fq′/Fm′ (= ФPSII′), is reduced under iron limitation  at each given light level, increased under iron limitation. We hypothesize that this is likely achieved by increased alternative electron transport pathways acting to keep RCIIs open (QA oxidized) and bypassing the electron flow bottleneck at PSI, when excitation pressure is high . In line with this explanation is the observation that values of σPSII are high under iron limitation, and rapidly decrease after iron addition . We observed a strong correlation between the conversion factor Φe:C/nPSII and the expression of NPQNSV .Above, we discussed how mechanisms acting down-stream of the initial charge separation in RCII are likely to be enhanced under conditions of high excitation pressure, resulting in high ETRing RCII . Fig 8 sing RCII –134. Foling RCII , we estif NPQNSV . We notee:C/nPSII and NPQNSV shown in e:C/nPSII and NPQNSV differentially, there are many processes related to the amount of excitation pressure experienced by the ETC that would influence both in a consistent manner. Numerous studies have shown that Фe:C increases if light is saturating, i.e. when excitation pressure is high . ,40. e:C/f NPQNSV .e:C/nPSII and NPQNSV  conversion factor, but are captured with our NPQNSV-based approach, as every single ETRRCII estimate is paired with a corresponding NPQNSV estimate. Finally, a non-static conversion factor is crucial if the goal is to monitor the effects of environmental change on marine primary productivity, since physiological responses to environmental change will likely affect the conversion factor itself before productivity changes are observed.While it remains to be seen how strong the correlation between Φd NPQNSV  may be fe:C/nPSIIvs. NPQNSV correlation determined from our iron addition experiment . Our approach consistently underestimates values from the deepest sampling depth, which can likely be attributed to the lack of spectral correction of our data. The RMSE for the values predicted using our approach and measured values is 48.4 mol C mol chl a-1 hr-1. This error represents ~ 10% of the total range of values observed along the transect during this study, suggesting that rates of productivity can be predicted with reasonable accuracy. In comparison with our approach, computation of CO2-assimilationfrom FRRF data assuming a constant 1/nPSII value of 500 mol chl a mol RCII-1and 4 mol e- mol C-1, significantly under-predicts observed CO2-assimilation rates (RMSE = 837.3 mol C mol chl a-1 hr-1). Even if we use a constant conversion factor derived from the average of the Φe:C/nPSII measured during our iron addition experiment, the model error remains larger compared to that derived using our variable, NPQNSV-based conversion factor . Future studies will be needed to evaluate the relationship between NPQNSV and Φe:C/nPSII in a number of oceanic regions in order to evaluate the potential for improved CO2-assimilation estimates from FRRF data.Crucial to this approach is a sound characterization of the conversion factor between FRRF-derived ETR regimes ,136. We S1 Fig14C-uptake experiments. (c) Spectral overlap of the two light sources. The overlap is good in the region of maximal light absorption by photosynthetic pigment (ca. 450 nm). However, in direct comparison with the photosynthetron, the FRRF instrument provides a higher proportion of photons in the region > 480 nm. This could have led to an underestimation of ETRRCII values relative to CO2-assimilation values measured in the photosynthetron, resulting in an under-estimate of Φe:C/nPSII. In addition to knowledge of spectral differences in the light sources used (a-c), spectral correction of our data would require light absorption spectra of the phytoplankton assemblages examined. Relative absorption spectra of the phytoplankton communities on day 6 after iron-addition .(a) The FRRF instrument used during this study contains LEDs with peak output at four wavelengths . In our FRRF instrument, excitation as well as actinic background irradiance is applied from the same LEDs. (b) Spectral distribution of the LEDs used in the photosynthetron used forechnique ) are sho(TIFF)Click here for additional data file.S2 Figa) was derived from HPLC analysis of accessory photosynthetic pigment. Average values are shown from three biological replicates for the iron-limited control and the iron addition treatment on day 6 of the experiment. One to1.5 L of water were filtered on 25 mm GF/F and stored at -80°C until analysis. Pigments were extracted and quantified as described by Taylor et al. [The taxonomic composition of phytoplankton assemblages Click here for additional data file.S3 FigvsE curves were fit with the exponential model of Webb et al. [The rates were measured as a function of irradiance, and Pb et al. . Shown a(TIFF)Click here for additional data file.


Lithium has a narrow therapeutic index with a subtle balance between effectiveness and adverse effects. Current guidelines recommend the use of lithium as a treatment for acute bipolar depression; however, the therapeutic range for the treatment has not been fully defined. Recently, the adjunctive lower lithium dose in bipolar depression has revealed potential efficacy; however, no study has investigated it predominantly in monotherapy. In this open-label, proof-of-concept study, 31 individuals with bipolar disorder during a depressive episode were randomized and 29 were followed up for six weeks with flexible lithium dosing. All subjects had a 21-item Hamilton Rating Scale for Depression (HAM-D) score of ≥18 at baseline. Subjects were divided into two groups, with higher (Li ≥0.5 mEq/l) or lower (Li <0.5 mEq/l) blood lithium levels. Response and remission rates were evaluated using the HAM-D scores. Following 6 weeks of lithium treatment, the remission rate for all patients was 62.0%. The plasma lithium levels did not impact the clinical response. However, subjects with higher blood lithium levels had an increased prevalence of nausea, restlessness, headaches and cognitive complaints. The results indicate that the lithium dose for the treatment of bipolar depression in an individual should be based on the clinical efficacy and side-effects. In the context of personalized psychiatric treatments, it is necessary to evaluate the therapeutic action of lithium with individual regimens in order to develop more tolerable and effective treatment approaches. Bipolar disorder (BD) is a chronic and disabling psychiatric disorder, with patients spending up to half of their time in a state of depression ,2. Thus,et al  . PatientMale and female outpatients, aged between 18–45 years were eligible to participate in the study. All subjects were recruited and followed up at the Mood Disorders Program, Laboratory of Neuroscience LIM27, Institute of Psychiatry of the University of São Paulo  between August 2010 and June 2012. Clinical assessments included: the score for the 21-item Hamilton Rating Scale for Depression (HAM-D) as the primary outcome, and the Young Mania Rating Scale (YMRS) and the Clinical Global Impression (CGI) scale scores as secondary outcomes. A HAM-D score of ≥18 was also required for inclusion. All adverse effects were recorded using the Udvalg for Kliniske Undersogelse (UKU) side-effects rating scale. Early improvement was defined as ≥20% improvement from the baseline HAM-D score following one week of lithium treatment. The response rate was defined as a reduction of ≥50% in the HAM-D score at endpoint and the remission rate was characterized as a HAM-D score of <8 at endpoint. There was no run-in period. At the onset of the study, all subjects had <3 mood episodes in their lifetime and an illness duration of ≤5 years. Exclusion criteria included: rapid cycling in the past 12 months, previous head trauma, a current Axis I psychiatric disorder other than BD, subjects submitted to electroconvulsive therapy, significantly abnormal laboratory test results and any chronic medical condition. The diagnosis was determined by experienced psychiatrists who held an inter-rater reliability score of >0.9 for HAM-D and YMRS. The first assessment was carried out at baseline [visit (V)1] and subsequent visits for clinical assessment, dose adjustment and monitoring of the levels of lithium were performed at the end of weeks 1 (V2), 2 (V3), 4 (V4) and 6 (V5). The present study was approved by the local institutional ethics committee of the Clinics Hospital  and all patients provided written consent prior to participation in the study.The primary outcome was the degree of change in depressive symptoms (HAM-D scores) analyzed using mixed-effects random regression. The mixed-effects regression was used to analyze changes in HAM-D assessments over the 6 weeks of follow-up and their association with time and clinical variables that were potentially associated with response to lithium treatment including: age, gender, duration of illness, history of psychosis, BD subtype and plasma lithium levels at week 6. Inter-group comparisons of continuous variables with a Gaussian distribution were performed using the Student’s t-test. The analyses of continuous variables with a non-parametric distribution were conducted using the Mann-Whitney U Test . The chi-square test was used for the comparison of categorical factors. The Pearson coefficient was used to evaluate the correlation between the relative change in HAM-D scores over time and other demographic and clinical variables. Statistical significance was set at P<0.05 (two-tailed) and data are presented as means ± standard deviations. All statistical analyses were carried out using SPSS software, version 16.0 .A total of 42 patients were accessed for eligibility to participate in the current study following a telephone screening at the Mood Disorders Program, LIM27, University of São Paulo. Of these, 31 patients were included in the present study. Two drop-outs were observed: one patient left due to personal problems unrelated to the study and the other patient was excluded due to noncompliance to the treatment (undetectable blood lithium levels). Thus, the final sample at endpoint comprised of 29 acutely depressed bipolar patients.The 29 patients who completed the study were followed-up with lithium monotherapy (n=25) or lithium therapy associated with another mood stabilizer or antipsychotics (n=4). The mean plasma level of lithium at V5 was 0.49±0.20 mEq/l.A significant reduction in HAM-D scores was observed over the six weeks of follow-up in the whole group of BD patients (P<0.001). Following six weeks of lithium treatment, the response and remission rates in the total study population were 86.2% and 62.0%, respectively. The mean HAM-D and YMRS scores following six weeks of lithium treatment were 7.3±5.9 and 3.8±8.6, respectively. When comparing BDI and BDII, similar remission rates were observed at endpoint; 63.6 vs. 61.1%, respectively . The CGIAt week 6, 15 patients had lower (Li<0.5 mEq/l) and 14 presented higher (Li≥0.5 mEq/l) plasma lithium levels. The plasma lithium levels did not differentially impact the number of patients who responded . Furthermore, the use of hypnotics (n=7), if necessary, was not observed to influence the clinical outcome . No other significant association was observed in the mixed-effects regression. No significant differences were observed in patients in remission (n=18) vs. those not in remission (n=11) at endpoint (a HAM-D score <8) with regard to oral dosage, blood lithium levels, gender and bipolar disorder subtype (data not shown). No association was observed between HAM-D scores and lithium dose (P=0.99) or blood levels (P=0.87) at endpoint.With regard to the potential predictors of response rate, only a previous history of psychosis was observed to be significantly associated with a faster reduction in HAM-D scores in response to lithium treatment . None of the patients experienced a switch to (hypo)mania during the follow-up period. Baseline YMRS scores did not influence the response rate (data not shown). Common adverse effects observed were polydipsia/polyuria (62.1%), cognitive complaints (41.4%), nausea (31.0%), increased oniric activity (31.0%) and sedation (31.0%). In subjects with higher lithium levels compared with those with lower levels, an increased prevalence of nausea , restlessness  and headaches  was observed. No patient drop-outs due to side-effects occurred during the follow-up period.In the present six-week, open-label, single-arm study, lithium revealed a significant antidepressant efficacy in short-term BD. No differences in efficacy were observed with lower levels (<0.5 mEq/l) vs. standard levels (0.5–1.2 mEq/l) of lithium. However, the current study population comprised predominantly young subjects with a shorter duration of illness and included predominantly BDII cases, which may have had an influence on the positive clinical outcome.et al did not identify a superior efficacy of using higher lithium levels (≥0.8 mm/l) in long-term maintenance therapy (The similarity in the efficacy of standard vs. ‘sub-therapeutic’ lithium levels that was observed in the BD treatment in the present study has been indirectly addressed by recent studies on BD ,16. The  therapy . Further therapy . Similar therapy . The limThe current study aimed to define the optimal lithium dose for BD treatment based on the clinical responses of patients and their tolerance of lithium. The results revealed that lower lithium doses demonstrated similar improvements to higher doses; however, they were associated with lower drop-out rates due to side-effects. In the context of personalized psychiatric treatments, it is critical to evaluate the therapeutic action of lithium with individual regimens in order to develop more effective and tolerable approaches to BD treatment. Further studies are required to investigate this.


A 16-year-old male patient was admitted to our hospital in March 2012 with chief complaints of systemic lymphadenopathy for >4 years with significant enlargement in the bilateral supraclavicular region for 1 month before admission. The patient had enlarged systemic lymph nodes, including the bilateral cervical, mediastinal and retroperitoneal lymph nodes, with no fever and night sweat initially in January 2008. Abdominal computed tomographic (CT) scan and ultrasonography revealed bilateral cervical, retroperitoneal lymphadenopathy and splenomegaly. The results of bone marrow cytomorphologic examination and biopsy were normal at that time. Biopsy of a left supraclavicular lymph node that was done in March 2008 at the first affiliated hospital of Zhengzhou University was suggestive of mixed-type Castleman's Disease. Enlarged lymph nodes significantly shrunk after 3 months of therapy with prednisone and interferon but they did not disappear. The patient had received CHOP-based regimen chemotherapy from June 2008. Lymphadenectasis was unpalpable and the splenomegaly was alleviated after treatment with six courses. Following this, the treatment was stopped after 6 months of interferon as maintenance therapy. Throughout the following 3 years, the patient was asymptomatic. However, in February 2012, he was admitted because of bilateral supraclavicular lymphadenopathy, which aggravated rapidly.β2-microglobulin and C-reactive protein (CRP) levels were 4.3 mg/l and 8.2 mg/l, respectively. Erythrocyte sedimentation rate was 38 mm in the first hour; anti-nuclear antibodies and rheumatoid factor were negative. Screening for human immunodeficiency virus, human herpes virus-8 (HHV-8), Epstein-Barr virus, and for hepatitis B and C viruses revealed negative results. All of the serum tumor markers tested  were negative. The thyroid function tests of T3, T4 and thyroid-stimulating hormone were normal. CT scans of the patient's chest and abdomen showed widespread lymphadenopathy in the thorax, axilla and retroperitoneal regions, as well as splenomegaly that was 14.9 cm × 4.7 cm in size. Electrocardiography test was normal.Physical examination revealed enlarged lymph nodes in the cervical, supraclavicular and axillary regions, the biggest of which was 6.4 cm × 3.9 cm in size in the left supraclavicular region. Splenomegaly  with no hepatomegaly was also palpated. Laboratory tests on admission showed a normal blood count. Urine analysis did not reveal protein traces . The serum interleukin(IL)-6 level was 32 pg/ml . No monoclonal immunoglobulin was detected in the serum by immunofixation electrophoresis. The serum Bone marrow biopsy was considered normal. Biopsy of a left supraclavicular lymph node revealed that the majority of germinal centers exhibited a tight/concentric pattern of follicular dendritic cell network, accompanied by vascular proliferation in the interfollicular space, and the typical onion-skin-like performance also appeared around germinal centers. . An immu2 as an intravenous bolus on days 1, 4, 8 and 11, repeated at 21–28 days for a total of eight cycles; the chemotherapy consisted of the standard hyper-CVAD program that was given alternating hyper-CVAD with high-dose methotrexate and cytarabine for six cycles every 21–28 days. The patient did not present with a serious peripheral polyneuropathy and four grade neutropenia. Partial remission was reached after two cycles of treatment. The serum IL-6, β2-microglobulin and CRP level decreased to 11.3 pg/ml, 2.8 mg/l and 3.2 mg/l, respectively. A 75% decrease in lymph nodes and splenomegaly was noted. The treatment was well tolerated, and the patient was in complete remission after the fourth cycle when evaluated with the positron emission tomography-CT scan. Bortezomib was administered for another two cycles after six courses of chemotherapy. After treatment, the patient was in good condition; lymphadenectasis and splenomegaly were unpalpable, and serum IL-6, β2-microglobulin and CRP levels decreased to 3.1 pg/ml, 1.9 mg/l and 2.4 mg/l, respectively. Thalidomide was administered on sleep at a dosage of 200 mg/day as maintenance therapy for 6 months after the treatment with bortezomib. Compared with pre-therapeutic levels, lymph nodes and spleen remained stable and no significant changes were noted, and the patient was asymptomatic at 24 months' follow-up.After obtaining signed informed consent from the patients, bortezomib combined Hyper-CVAD  routine chemotherapy was initiated. Bortezomib was given at the standard dose of 1.3 mg/m1 Progressive research on the pathogenesis of Castleman's disease has identified some novel agents including anti-IL-6 monoclonal antibody, humanized anti-human IL-6 receptor monoclonal antibody and rituximab for the MCD;4 a small number of MCD patients who were administrated bortezomib were also reported.5Multicentric Castleman's disease (MCD) is a subtype of Castleman's disease. It has been reported as a rare benign disease characterized by lymphocyte proliferation originally. Common therapies including steroid monotherapy and combined low-dose chemotherapy have been widely used, but there continue to be a number of patients in whom treatment is ineffective or in whom relapse occurs rapidly after the termination of treatment; in particular, the plasma cell subtype always has a highly aggressive feature.6 we decided to treat this patient with bortezomib associated with Hyper-CVAD chemotherapy. Clinical improvement was achieved after completion of the second cycle of treatment. The patient was well tolerant to the treatment and showed a definitive clinical improvement and sustained reduction of IL-6, β2-microglobulin and CRP levels.The patient in our study presented with progressing systemic lymphadenectasis at first, followed by significant bilateral supraclavicular lymph node enlargement. Immunohistochemistry and pathology of the left supraclavicular lymph node revealed the diagnosis of mixed-type Castleman's Disease in March 2008. After the common chemotherapy, lymphadenectasis was unpalpable and splenomegaly was alleviated. His condition was stable with no recurrence until February 2012. However, in February 2012, 3 years after the end of treatment, this patient relapsed. The same histopathology result was confirmed by the biopsy of the left supraclavicular lymph node. However, a more rapid progression of enlarged lymph nodes compared with 3 years ago was observed. To our knowledge, there is still no consensus treatment guideline for the relapsed MCD; obviously, it was difficult to achieve the desired effect for this relapsed patient through conventional chemotherapy. It was considered that the patient had a mixed hyaline-vascular and plasma cell type of MCD. Bortezomib has been reported to inhibit myeloma cell growth by NF-κB blockade and decrease IL-6 level. On the basis of the efficacy of bortezomib against B-cell malignancies and the experience in MCD reported in recent years,10 For this patient, in order to consolidate the effect, thalidomide was applied for 6 months as maintenance therapy. No treatment-related side effects occurred and the patient showed persistent clinical improvement.As is well known, thalidomide also has similar immunomodulatory and anti-angiogenetic properties to INF-alpha. in addition, thalidomide may decrease the serum IL-6 levels. Some studies reported that bortezomib and thalidomide were effective in the treatment of Castleman's Disease.Of note, in our study, administration of bortezomib-combined conventional chemotherapy followed by thalidomide successfully improved the condition of the patient with MCD, and induced significant and durable responses. Therefore, it is suggested that bortezomib and thalidomide could be used as salvage therapy for refractory MCD.


LPS caused a dose-dependent increase of secretion of IL-8 by cultured UM. Lutein and zeaxanthin did not affect the constitutive secretion of IL-8. However, lutein and zeaxanthin decreased LPS-induced secretion of IL-8 in cultured UM in a dose-dependent manner. LPS significantly increased NF-κB levels in cell nuclear extracts and p-JNK levels in the cell lysates from UM, but not p-p38 MAPK and p-ERG. Lutein or zeaxanthin significantly reduced LPS-induced increase of NF-κB and p-JNK levels, but not p38 MAPK and ERG levels. The present study demonstrated that lutein and zeaxanthin inhibited LPS-induced secretion of IL-8 in cultured UM via JNK and NF-κB signal pathways. The anti-inflammatory effects of lutein and zeaxanthin might be explored as a therapeutic approach in the management of uveitis and other inflammatory diseases of the eye.The effects of lutein and zeaxanthin on lipopolysaccharide- (LPS-) induced secretion of IL-8 by uveal melanocytes (UM) were tested in cultured human UM. MTT assay revealed that LPS (0.01–1  Zeaxanthin and lutein are two natural bioactives that belong to the xanthophyll class. They are present in the eye in high concentrations. The retina and lens in general and the macular region in the center of the retina in particular are highly enriched in these two xanthophylls , 2. TherLPS is an endotoxin that can induce a potent inflammatory response . InjectiAnimal studies indicated that lutein reduces the secretion of proinflammatory cytokines and chemokines and this may be the mechanism of inhibition of LPS-induced uveitis by lutein . HoweverUveal melanocytes (UM) are the predominant cell type in the uvea. In the past, very little was known about the function of UM and the role of UM in the pathogenesis of various eye diseases. In the past decades, after the development of methods for the culture of UM and the establishment of in vitro models for studying the function of UM, it has been reported that UM produce various growth factors, cytokines, and chemokines, in addition to their functions related to melanin –19. Hu eκB) ELISA kits were obtained from Invitrogen .Cell culture medium, fetal bovine serum, and trypsin were obtained from GIBCO . LPS, dimethyl sulfoxide (DMSO), collagenase, lutein, 3--2,5-diphenyltetrazolium bromide (MTT), phosphate buffered saline (PBF), and protease inhibitor cocktail were obtained from Sigma . Zeaxanthin was obtained from ZeaVision LLC . Quantikine IL-8 ELISA kit was obtained from R&D System . p38 mitogen-activated protein kinase (MAPK), extracellular signal-regulated kinases 1/2 (ERK1/2), and c-Jun N-terminal kinase 1/2 (JNK1/2) ELISA kits and cell extraction buffer, PMSF, hypotonic buffer, and nuclear factor-kappa B .MTT assay was used to determine the effects of LPS, lutein, and zeaxanthin on cell viability of cultured UM as previously described . For eac5 cells/well) were seeded into the 24-well plates and cultured. When cells reached 80–90% confluence, they were rinsed with PBS and incubated in serum-free culture medium with or without LPS. After 24 h, the conditioned media were collected and centrifuged. The supernatants were stored at −80°C.Human UM (1 × 105 cells/well) were seeded into the 24-well plates and cultured. When cells reached 80–90% confluence, they were rinsed with PBS and incubated in serum-free culture medium with or without lutein or zeaxanthin. After 24 h, the conditioned media were collected and centrifuged. The supernatants were stored at −80°C.Human UM (1 × 105 cells/well) were seeded into the 24-well plates and cultured, and culture medium was replaced as described above. Lutein or zeaxanthin at different final levels was added. Two hours later, LPS at the final levels of 0.1 μg/mL was added to the cultures. After 24 h, the conditioned media were collected and centrifuged. The supernatants were stored at −80°C.Human UM (1 × 10Enzyme-linked immunosorbent assay (ELISA) was used for the measurement of IL-8 protein levels in the supernatant of cultured cells. A commercially available Quantikine IL-8 ELISA kit was used to determine IL-8 protein levels according to the protocol provided by the manufacturer. The optical density of the ELISA samples was measured at 450 and 540 nm using a microplate reader. The sensitivity of the assay for IL-8 was 3.5 pg/mL.6 cells/well) were seeded into 6-well plates and cultured for 24 h and culture medium was replaced as described above. Lutein or zeaxanthin at different final levels was added. Two hours later, LPS at the final levels of 0.1 μg/mL was added into cultures and cultured for 60 min. The cultured media were removed and the cultures were washed with PBS. UM were harvested by scraping with a rubber policeman. Cell were washed with PBS and centrifuged. Cell extraction buffer with protease inhibitor cocktail and PMSF were added to the pellets and cultured for 30 min and centrifuged. All of these procedures were done under 4°C. The supernatants were collected and stored at −80°C. For the study of NF-κB, cells were cultured, treated with or without lutein, zeaxanthin, and LPS, and collected as described above. Collected cells were treated with hypotonic buffer and centrifuged. The pellets that contain nuclear fraction were collected, treated with cell extraction buffer, vortexed, and centrifuged. The supernatants were stored at −80°C.For the study of phosphorylated- (p-) p38 MAPK, p-ERK1/2, and p-JNK1/2 levels, human cultured UM .Each experiment was replicated 3 times and the data were presented as mean ± standard deviation (SD). A one-way analysis of variance (ANOVA) test was performed to assess the significance. Values of μg/mL did not influence the cell viability of cultured human UM    . Lutein  Figures . Therefoμg/mL caused a dose-dependent significant increase of IL-8 levels , indicating a low level of constitutive secretion of cultured UM. LPS at 0.01–1.0 μM) did not significantly differ from those from cells not treated with lutein and zeaxanthin  dose-dependently decreased LPS-induced secretion of IL-8 in cultured UM , suggesting that lutein has a partial inhibitory effect on LPS-induced secretion of IL-8. Zeaxanthin  dose-dependently decreased LPS-induced secretion of IL-8 in cultured UM , suggesting that zeaxanthin also has a partial inhibitory effect on LPS-induced secretion of IL-8.Lutein  play an important role in LPS-induced increased secretion of IL-8 and in the inhibitory effects of lutein and zeaxanthin on LPS-induced increased secretion of IL-8.LPS at 0.1 κB signaling pathways.In the present study, we demonstrated that lutein and zeaxanthin inhibited LPS-induced secretion of IL-8 in cultured human UM and this effect was mediated by JNK1/2 and NF-Uveitis is a common eye disease and a major cause of visual impairment throughout the world , 11, 20.LPS is an endotoxin and is the major component of the outer membrane of Gram-negative bacteria. LPS can induce a strong response from the immune system . LPE-indChemokines act as chemoattractants and activators of specific leukocytes at the site of inflammation. Chemokines could be classified into four subfamilies based on the number and location of the cysteine residues at the N-terminus of the molecule and are named CC (with two adjacent cysteines near the N-terminus of the molecule), CXC (the two cysteines being separated by an amino acid), CX3C (having three amino acids between the two cysteines), and C (having a specific amino acid sequence of glutamic acid-leucine-arginine immediately before the first cysteine), in agreement with the systematic nomenclature. In the two main subfamilies (CC and CXC), CXC chemokines are important in the attraction of neutrophils and CC chemokines have powerful chemoattractants and activators for monocytes and lymphocytes , 21, 22.It has been reported that intravenous injection or oral administration of lutein suppresses the development of LPE-induced uveitis in rats and mice, respectively , 11. TheIn the present study, LPS significantly induced the secretion of IL-8 by UM in a dose-dependent manner, which is consistent with previous reports . Lutein κB signal pathway, but not p38 and ERK pathway; this is consistent with the animal study, which showed that lutein inhibited the activation of NF-κB in the iris-ciliary body in LPS-induced uveitis and in cultured macrophages [The mechanism of lutein and zeaxanthin inhibition of LPS-induced expression of chemokines in UM has not been previously reported. The present study demonstrated that lutein and zeaxanthin inhibited the secretion of IL-8 induced by LPS through the activation of JNK1/2 and NF-rophages , 10.κB signaling pathways. Inhibition of secretion of IL-8 by lutein and zeaxanthin might be explored as a therapeutic approach in the management of uveitis and other inflammatory diseases of the eye.In conclusion, this study demonstrated that lutein and zeaxanthin inhibited LPS-induced secretion of IL-8 in cultured human UM and this effect was mediated by JNK1/2 and NF-


Si-based integrated circuits have been intensively developed over the past several decades through ultimate device scaling. However, the Si technology has reached the physical limitations of the scaling. These limitations have fuelled the search for alternative active materials (for transistors) and the introduction of optical interconnects . A series of attempts to circumvent the Si technology limits are based on the use of III-V compound semiconductor due to their superior benefits, such as high electron mobility and direct bandgap. To use their physical properties on a Si platform, the formation of high-quality III-V films on the Si (III-V/Si) is the basic technology ; however, implementing this technology using a high-throughput process is not easy. Here, we report new concepts for an ultra-high-throughput heterogeneous integration of high-quality III-V films on the Si using the wafer bonding and epitaxial lift off (ELO) technique. We describe the ultra-fast ELO and also the re-use of the III-V donor wafer after III-V/Si formation. These approaches provide an ultra-high-throughput fabrication of III-V/Si substrates with a high-quality film, which leads to a dramatic cost reduction. As proof-of-concept devices, this paper demonstrates GaAs-based high electron mobility transistors (HEMTs), solar cells, and hetero-junction phototransistors on Si substrates. Historically, III-V compound semiconductors have been explored as active materials for high-speed electronic devices13578101214Recently, high-quality III-V/Si formation method based on wafer bonding have been rapidly developed by many research groups, with the successful production of high-quality III-V films on Si substrates516171819212217et al. first proposed the method27Another approach to wafer re-use is the use of epitaxial lift off (ELO) techniques. ELO was developed in the 1980s and 1990s25In this work, we describe an ultra-fast ELO process that involves a pre-patterning step before the wafer bonding and the use of etching acceleration solutions. Furthermore, we demonstrate that the III-V donor wafer can be re-used after the wafer bonding and ELO process at least once. These approaches enable a high-throughput fabrication of GaAs/Si substrate with a high-quality film of GaAs, which lead to a dramatic cost reduction, as estimated in 0.85Ga0.15As sacrificial layer to protect the active layer and the GaAs donor wafer during the ELO process. The main cause of the long processing time for the ELO was known to be the formation of H2 bubbles and the etching residues generated during the ELO process28292 bubble release and increase the accessible etching areas, a pre-patterning step that breaks the III-V layer into smaller pieces was added between the growth of III-V layer and the wafer bonding. Actually, the use of the patterned channel was reported for the outgassing in III-V/Si hybrid structure, which enhanced thermal stability due to efficient gas release through the channels3132330.85Ga0.15As sacrificial layer in the ELO process. As we explained afterwards, etching acceleration solutions were isopropanol (IPA) and acetone (Ace), which accelerate the etching of Al0.85Ga0.15As sacrificial layer by making the H2 gas release smoothly and reducing the etch residues. Here, one possible concern is the surface contamination during the patterning and etching. Therefore, for the high yield bonding, the careful surface preparation before the wafer bonding is needed. In our experiment, overall transfer yield was higher than 95%. Next, the wafer bonding was conducted at room temperature, followed by the device fabrication  solutions, which directly shows the difficulty of the use of a conventional ELO process. With a pre-patterning step, the ELO time was significantly reduced to 6 hours in the same solutions. This reduction is attributed to the increase of an exposed surface area of an Al0.85Ga0.15As layer due to the pre-patterning, which enabled the efficient gas bubbles release3132332 bubbles from becoming large. This process promotes the rapid release of H2 bubbles from the samples. With increasing HF concentration and the addition of etching acceleration solutions, the ELO time was significantly reduced down to approximately 20 min. Comparing to that of other study with a channel release of 110 min, the ELO time achieved in this study was quite short293031323334To investigate the effect of the pre-patterning step and the etching acceleration solutions, we compared the ELO process for various etching conditions. rms) of 0.02 nm. After the ELO process, the Rrms of the surface was increased up to 0.5 nm and etching residues with dot shapes were observed over the entire wafer. After the cleaning using HCl and the etching of the etch stop layer, the AFM images illustrate the excellent surface morphology that is comparable to that of the epi-ready surface. The AFM image of the surface after re-growing the GaAs layer confirmed a high process stability with a flat and smooth surface. To investigate the layer quality and the possible strain in the transferred film, we collected Raman spectra, as shown in 270.85Ga0.15As sacrificial layer between a device layer and the GaAs donor wafer -gate voltage (VG) characteristics of the GaAs-OI HEMT with a gate length (LG) of 2 μm. Clear transfer curves were obtained with a steep subthreshold slope (S.S. = dVG/d[log(ID)]) of 83 mV per decade and high on/off ratio (Ion/Ioff) of 107. To the best of our knowledge, the achieved S.S. and Ion/Ioff are record values among the reported III-V-OI transistors17181920Ioff was found to be a very low value of <1 pA/μm, which is lower than that required for an ultra-low power (ULP) transistor (10 pA/μm) by the International technology roadmap for semiconductors (ITRS). The clear current saturation was also observed in the ID-drain voltage (VD) characteristics  of the Si substrate. Since GaAs channel layer and underlying Si substrate are electrically isolated, a channel potential can be modulated via VB biasing without a leakage current. ID-VG curves with changing VB from 0.5 to −0.5 V. The ID-VG curves were intentionally shifted with changing VB, indicating that the device characteristics can be controlled after the device fabrication, which provides additional functionality for a circuit design.eristics . From thNs) of fresh and re-grown GaAs HEMTs at 80 K and 300 K. The mobility and Ns behaviors were almost the same between the two sets of devices, which indicates the high quality of the re-grown GaAs HEMT layer, whereas the Raman spectra was slightly broader. These results strongly suggest that the epi-ready surface of the GaAs donor wafer can be recovered after the ELO process, thereby enabling the donor wafer to be re-used at least once. Further development will be expected via the optimization of the surface treatment condition before the re-growth with a quality evaluation by the Raman spectra, Hall mobility, and each device performance.Even though the re-grown GaAs layer on the donor wafer after the ELO process was evaluated by AFM and Raman spectra, these results do not guarantee the re-use of the donor wafer because these measurements do not fully reflect the epitaxial quality. Here, the best way to show the wafer-reusability is to fabricate and compare each types of device. However, for the simple examination, Hall mobility is the one of the best figure-of-merit to evaluate the crystal quality of grown epitaxial film, which is typically used to check/confirm the status of the growth chamber. Therefore, we have investigated the electrical properties of re-grown HEMTs. 0.85Ga0.15As sacrificial layer, followed by Pt/Au deposition  formation and etching of the GaAs top contact layer  – V curves of the GaAs solar cell on Si under air mass (AM) 1.5 G, 1-sun measurement conditions. As a control device, the data of a GaAs solar cell grown on GaAs is also shown. Even considering approximately 30% reflection of the incident light due to the absence of an anti-reflection coating, the energy conversion efficiency (η) and the open-circuit voltage (Voc) of the solar cell on Si was as high as 14.05% and 0.89 V, respectively. The performance of the solar cell on Si was almost same as that of the control device with an even higher short-circuit current (Jsc). The external quantum efficiency (EQE) of both devices in Jsc value is different between the two. We believe that light reflection from the back side enhances photo recycling of the light once reached to the back-side, leads to higher Jsc in the solar cell on Si. However, since the intensity of an incident light to the sample was very small in EQE measurement, the data could not catch the light response reflected from back side metal. These results strongly suggest that the developed wafer bonding and ultra-fast ELO techniques provide a high-quality III-V film on Si substrate without material degradation during these processes.VCE). Pin) of 1.2 μW at a wavelength of 635 nm. The data from the HPTs fabricated by both the developed ultra-fast ELO and the conventional ELO without the pre-patterning and accelerant addition are shown. For both devices, −13 A over the entire VCE range. This low ICdark confirms the defect-free crystal quality after the wafer bonding and ELO processes. In contrast to the VCE > 0.25 V, Gopt = hυΔIc/qPin, where hυ and ΔIc are the incident photon energy and the difference between Gopt of the HPT fabricated using the ultra-fast ELO process shows higher values, demonstrating another benefit of the developed ELO process besides the time reduction. Furthermore, array formation with each HPTs using the developed process can be expanded to the focal plane array for the high resolutionGaAs HPTs on Si were also fabricated using similar procedures. After growing an epitaxial layer comprising a GaAs collector, a GaAs base, and an InGaP emitter, the III-V and Si wafers were bonded to each other using a Pt/Au bonding layer, followed by the ELO  with AsH3, PH3, TMGa, TMIn, and TMAl as the precursors. SiH4 and DMZn were used for n- and p-type dopants. The GaAs solar cells were composed of a p-type GaAs contact layer, and a p-type InGaP back surface field layer, a p-type GaAs base, an n-type GaAs emitter, an n-type GaAs window layer, and n-type GaAs contact layer. For the ELO process, an Al0.85Ga0.15As sacrificial layer was inserted between the device layer and the GaAs donor wafer. Optionally, an etch stop layer of InGaP/GaAs was also inserted between the Al0.85Ga0.15As layer and the device layer and also between the Al0.85Ga0.15As layer and the GaAs donor wafer. To check the quality of the epitaxial film, Hall measurement was carried out with 0.7 × 0.7 cm2 pieces at 0.55 T magnetic field using HMS 3000 system, Ecopia. During the measurement, no bias gating was applied.A 2-inch GaAs wafer (001) was prepared for an epitaxial growth for all experiments. The epitaxial layers of the GaAs HEMTs and InGaP/GaAs HPTs were grown by solid source molecular beam epitaxy (MBE) with the use of Si and Be cell as sources of n- and p-type dopants. GaAs HEMTs were composed of an n-type contact layer, an Al4OH and HF solutions, respectively, to remove the native oxides. Next, bonding materials, i.e., Y2O3 or Pt/Au in this study, were deposited via electron beam evaporation. A pre-patterning process was performed using H3PO4- and HCl-based solutions for the etching of Al(Ga)As and InGaP, respectively. Subsequently, the wafers were cleaned by acetone in an ultra-sonic bath, followed by surface plasma treatment using O2 and Ar for Y2O3 and Pt/Au, respectively. The surface-treated wafers were bonded to each other in the air, followed by uniaxial pressing with a force of 180 kgf. The ELO process was performed in HF-based solutions until the separation of the III-V/Si and the GaAs donor wafer was completed. The completion of ELO was determined when III-V/Si wafer and III-V donor wafer were separated. Here, we observed the sample separation on every 1 min after dipping the samples in HF-based solutions.III-V and Si wafers were cleaned by NH3PO4-based solution, followed by Y2O3 field-oxide deposition. S/D contacts were formed by Pd/Ge/Au deposition and rapid thermal annealing (RTA) at 150 °C for 3 hours. Next, the contact layer was selectively etched by citric acid:H2O2 (3:1 volume ratio) solutions for the gate electrode formation. Finally, the gate electrode of Ti/Pt/Au was deposited. Each device was characterized using a semiconductor parameter analyser  in the dark.Mesa etching was performed using a H3PO4-based solutions through the top electrode as a mask. Each device was characterized using a solar simulator , IPCE .After the wafer bonding with a Pt/Au bonding material, the Pt/Au layer acted as a bottom electrode for the GaAs solar cells. The top electrode of Ni/Au/Ge/Ni/Au was deposited, followed by RTA at 400° C for 40 sec. Next, the n-type GaAs contact layer was selectively etched using H3PO4-based solutions through the top electrode as a mask. The optical performances of the fabricated devices were characterized utilizing a 635-nm laser diode and a semiconductor parameter analyser . For laser power calibration, the incident optical power through a lensed fiber was measured using an optical meter  equipped with a silicon photodetector  in the dark.Similar to the GaAs solar cells, the Pt/Au layer acted as an emitter electrode (located at the bottom) after the ELO process. The top electrode of Ni/Au/Ge/Ni/Au was deposited, followed by RTA at 400 °C for 40 sec. Next, the n-type GaAs collector contact layer was selectively etched using HHow to cite this article: Geum, D.-M. et al. Ultra-high-throughput Production of III-V/Si Wafer for Electronic and Photonic Applications. Sci. Rep.6, 20610; doi: 10.1038/srep20610 (2016).